DESCRIPTION: Ocular disease caused by herpes simplex virus affects up to 500,000 people in the United States. It is known that innate host resistance, host immune responses, and viral genes affect the severity of ocular disease. Several viral genes that affect virulence are known, but their role in ocular infection is not known. In addition, multiple viral genes likely interact to promote virulence and little is known about such interactions and how they affect virulence. Previously, it was shown that mixed ocular infection with two HSV-1 strains (OD4 and CJ394) resulted in a synergistic increase in both ocular disease and neurovirulence. The overall goal of the project is to identify the virulence determinants in CJ394 and OD4 and study their role in ocular disease. Marker transfer studies have mapped one determinant that transfers increased ocular disease to a 1.9 Kb region of the HSV-1 genome that contains part of the unique short (US) region of the genome (US1/1.5 genes) and a portion of the repeated regions flanking US. Sequencing a portion of the 1.9 Kb region has identified a frameshift/premature termination mutation in OD4 that would truncate the US1 ((22) protein at amino acid 68. The focus of the proposal is to confirm this represents one virulence determinant. At least two other regions of the CJ394 genome also transfer partial ocular virulence, suggesting multiple genes cooperate in increasing ocular disease. The Specific Aims are to (1) genotypically confirm that the OD4 US1 gene is mutated compared to CJ394, (2) phenotypically confirm that (22 functions are missing or compromised in OD4 but are normal in CJ394, (3) localize the virulence determinants in the other CJ394 fragments (EcoRI O and D), and (4) test the hypothesis that multiple virulence determinants cooperate to increase ocular virulence. This work will clarify the interactions between virulence genes, discover new virulence genes, and provide information that will improve therapy for ocular HSV infections.